Binding of platelet-derived growth factor (PDGF) to its cell surface receptors stimulates a variety of biochemical and biological responses. Two distinct PDGF receptors have been identified and each exhibits different binding affinities for the variant PDGF isoforms. The alphaPDGFR binds PDGF-AA and PDGF-BB, whereas the betaPDGFR binds PDGF-BB only. In NIH/3T3 cells where both PDGFR subtypes are coexpressed, the higher transforming function of PDGF-BB as compared to that induced by PDGF-AA was shown to be directly linked to distinct substrate specificity of betaPDGFR kinase. Construction and characterization of chimeric molecules between alphaPDGFR and betaPDGFR defined a domain responsible for higher transforming activity of PDGF-BB mediated by betaPDGFR kinase. Construction and characterization of deletion mutants of alphaPDGFR defined a domain responsible for PDGF-A mediated transformation in NIH/3T3 fibroblasts. Binding of PDGF to its cell surface receptor results in tyrosine phosphorylation of a number of cellular proteins which are thought to be critical for receptor-mediated mitogenic response in vivo. In an attempt to identify novel signaling molecules, a polyclonal antibody was generated against a partially purified substrate of PDGFR kinase domain. this antibody was used to identify clones which have been characterized to include some novel substrates.